OJIP fluorescence kinetics is widely used in the study of algae photosynthesis, but in common OJIP curve analysis, the characteristic time of J and I points is often fixed. Therefore, the influence of algae species on the characteristic time of J and I points is ignored, which results in the deviation of the fluorescence intensity calculation results of J and I points and directly affects the accuracy of the measurement results. In view of this, a method of using three-level exponential function to approximate OJIP curve is proposed to dynamically obtain the characteristic time of J and I points. The test results of experiments of characteristic time of J and I points of different algae species, and the characteristic time of J point under dichlorophenyl dimethylurea (DCMU) stress show that the proposed method can effectively obtain the characteristic time of J and I points of different algae. The characteristic time of J point of Scenedesmus dimorpha, Chlorella pyrenoidosa, Chlorella vulgaris and Cylindrotheca closterium is 2.22, 1.52, 1.33, 1.01 ms, respectively. The corresponding characteristic time of I point is 28.80, 27.15, 29.90, 15.28 ms, respectively. The relative standard deviation of multiple calculation results is less than 10%, which has a great consistency. Under the toxic stress test conditions with DCMU concentrations (mass concentrations) of 10, 20, 40 μg/L, the characteristic time of J point of Chlorella vulgaris calculated by the proposed method is 1.25, 1.18, and 1.10 ms, respectively, and the relative standard deviation is 12.03%. In addition, the photosynthetic activity parameter V_J calculated by the proposed method has a good toxic dose-effect relationship with the DCMU concentration, and the correlation coefficient R² is 0.991.